Mesothelial cells have been found to be pivotal in tumor metastasis, peritoneal dialysis, and inflammatory response. These cells are specialized epithelial cells that line the peritoneal, pericardial, and pleural cavities and internal organs presenting a barrier to invading organisms and physical damage. Ovarian tumor attachments occur through cancer cells binding to mesothelial cells and migrating into the surrounding tissue and vasculature. Peritoneal dialysis relies on the intact transport function of mesothelial cells to allow transfer of waste products from the underlying vasculature to the dialysis fluid in the peritoneal cavity. Host response to peritoneal infection is mediated by the inflammatory cascade initiated and cytokines release by peritoneal mesothelial cells. Research into the biology of mesothelial cells continues with the goal of developing methods to provide more effective treatments for disease. Currently, there are no commercial sources of these cells in the U.S. which requires researchers to isolate and characterize the cells on their own. This process is time consuming, requires access to human tissue, and introduces variation in the preparation and quality of the cells. Zen-Bio will address this need by providing a well characterized mesothelial cell culture system to the research community. Phase I of this project focuses solely on generating an optimized primary human mesothelial cell system. The Specific Aim is to optimize the isolation and propagation of human omental mesothelial cells followed by careful characterization of the cellular properties. Isolation and propagation optimization will be achieved by systematically modifying aspects of existing protocols to enhance viable cell yields while minimizing population doublings. A detailed analysis of cell properties, including longevity in culture, cell specific gene expression, and cytokeratin production will be performed to validate the cell system. These analyses will form the basis of future quality control procedures. Phase II of this project contains two Specific Aims that will generate additional contract services at Zen-Bio. Aim I is to analyze the cytokine and gene expression changes during mesothelial cell transdifferentiation (epithelial to mesenchymal transition). Completion of this Aim leads directly to Aim II which is to establish mesothelial cell contract services at Zen-Bio. These services include analysis of cytokine secretion, gene expression, tumor cell-mesothelial cell interaction, and cytotoxicity. Multiple product offerings are expected from this proposal: a primary human mesothelial cell system, support media and reagents, contract assay services, and kits for mesothelial cell research. At the completion of this project, a commercially available, fully characterized primary human mesothelial cell system and support reagents will be offered to researchers. The accessibility of this currently unavailable system will provide wider opportunity to investigate novel methods to inhibit ovarian tumor attachment, prolong the utility of peritoneal dialysis, and treat peritonitis. [unreadable] [unreadable] [unreadable]